目的 基于NLRP3炎症小体探讨黄芪赤风汤（HQCF）通过改善动脉粥样硬化（AS）内皮损伤的分子机制。方法 氧化低密度脂蛋白（ox-LDL）诱导bEnd.3细胞构建内皮细胞损伤模型，结合si-激活NOD样受体家族含pyrin结构域蛋白3（NLRP3）基因敲低技术及HQCF (50、100、200μg·mL^-1)干预。乳酸脱氢酶（LDH）试剂盒检测bEnd.3细胞受损伤程度；试剂盒法检测bEnd.3细胞内总胆固醇（TC）、游离胆固醇（FC）、胆固醇酯（CE）水平和CE/TC；实时荧光定量PCR（qRTPCR）检测bEnd.3细胞炎性因子白细胞介素-1β（IL-1β）、IL-6、IL-18和肿瘤坏死因子-α（TNF-α）和黏附分子细胞间黏附分子-1（ICAM-1）、血管细胞黏附分子-1（VCAM-1）基因表达水平；DCFH-DA探针通过流式细胞仪检测bEnd.3细胞内活性氧（ROS）含量；Western blotting检测bEnd.3细胞中Toll样受体4（TLR4）、p-核因子κB（NF-κB）/NF-κB、核苷酸结合寡聚化结构域样受体蛋白3（NLRP3）、含半胱氨酸的天冬氨酸蛋白水解酶-1（Caspase-1）和凋亡相关斑点样蛋白（ASC）蛋白表达水平；qRT-PCR评估si-NLRP3序列的转染效率；油红O染色评估b End.3细胞内脂质聚积情况。结果 与模型组相比，HQCF组LDH释放量显著减少（P<0.05、0.01）,TC、FC、CE水平及CE/TC显著降低（P<0.05、0.01）,ROS水平显著下调（P<0.01）,IL-1β、IL-6、IL-18、TNF-α m RNA表达水平显著下调（P<0.01）,TLR4、p-NF-κB/NF-κB、NLRP3、Caspase-1和ASC蛋白表达水平显著下调（P<0.01）。与ox-LDL+si-NC组相比，ox-LDL+si-NLRP3组和ox-LDL+si-NLRP3+HQCF组细胞脂质蓄积情况明显改善，且以ox-LDL+si-NLRP3+HQCF组改善效果最佳；ox-LDL+si-NLRP3组细胞TC和CE水平显著下调（P<0.01）,FC和CE/TC有下调趋势但无统计学意义，ox-LDL+si-NLRP3+HQCF组细胞TC、FC、CE、CE/TC的水平显著下调（P<0.01）;ox-LDL+si-NLRP3组与ox-LDL+si-NLRP3+HQCF组bEnd.3细胞ICAM-1和VCAM-1 mRNA表达水平显著下调（P<0.01）。结论 HQCF通过抑制NLRP3炎症小体通路激活，减轻ox-LDL诱导的内皮细胞脂质蓄积及炎症，从而发挥抗AS内皮细胞损伤的作用。

Objective To explore the molecular mechanism of Huangqi Chifeng Tang（HQCF） in improving endothelial damage of atherosclerosis（AS） based on NLRP3 inflammatory inflammasome. Methods ox-LDL-induced bEnd.3 cells were used to construct an endothelial cell injury model, combined with si-NLRP3 gene knockdown technology and HQCF(50, 100, 200 μg·mL^-1) intervention. Lactate dehydrogenase（LDH） assay kit was used to detect bEnd.3 cell viability; The assay kit was used to detect the levels of total cholesterol（TC）, free cholesterol（FC）, cholesterol esters（CE）, and CE/TC in bEnd.3 cells; Real-time fluorescence quantitative PCR（qRT-PCR） was used to detect the gene expression levels of inflammatory factors interleukin-1β（IL-1β）, IL-6, IL-18, tumor necrosis factor-α（TNF-α）, intercellular adhesion molecule-1（ICAM-1）, and vascular cell adhesion molecule-1（VCAM-1） in bEnd.3 cells; DCFH-DA probe was used to detect the intracellular reactive oxygen species（ROS） content in bEnd.3 cells by flow cytometry; Western blotting was used to detect the protein expression levels of Toll like receptor 4（TLR4）, p-nuclear factor kappa B（p-NF-κB）/NF-κB, NOD-, LRR-and pyrin domain-containing protein 3（NLRP3）, Cysteinyl aspartate specific proteinase-1（Caspase-1）, and Apoptosis-associated speck-like protein containing a caspase recruitment domain（ASC） in bEnd.3 cells; transfection efficiency of the si-NLRP3 sequence was evaluated by qRT-PCR; Oil red O staining was used to evaluate the intracellular lipid accumulation in bEnd.3 cells. Results Compared with model group, the LDH release in the HQCF group was significantly reduced（P＜0.05, 0.01）, the levels of TC, FC, CE and CE/TC were significantly decreased（P＜0.05, 0.01）, the ROS level was significantly down-regulated（P＜0.01）, the mRNA expression levels of IL-1β, IL-6, IL-18 and TNF-α were significantly down-regulated（P＜0.01）, and the protein expression levels of TLR4, p-NF-κB/NF-κB, NLRP3, Caspase-1 and ASC were significantly down-regulated（P＜0.01）. Compared with the ox-LDL + si-NC group, the lipid accumulation in cells of the ox-LDL + si-NLRP3 group and the ox-LDL + si-NLRP3 + HQCF group was significantly improved, and the improvement effect was the best in the ox-LDL + si-NLRP3 + HQCF group; the levels of TC and CE in cells of the ox-LDL + si-NLRP3 group were significantly down-regulated（P＜0.01）, FC and CE/TC showed a downward trend but without statistical significance, and the levels of TC, FC, CE and CE/TC in cells of the ox-LDL + si-NLRP3 + HQCF group were significantly down-regulated（P＜0.01）; the mRNA expression levels of ICAM-1 and VCAM-1 in bEnd.3 cells of the ox-LDL + si-NLRP3 group and the ox-LDL + si-NLRP3 + HQCF group were significantly down-regulated（P＜0.01）. Conclusion HQCF exerts an anti-AS endothelial cell injury effect by inhibiting the activation of NLRP3 inflammasome pathway, reducing ox-LDL induced lipid accumulation and inflammation in endothelial cells.

R285.5

黑龙江省自然科学基金项目(PL2024H239)